RESUMO
There have been few reports of vascularized fibular transfer for septic arthritis of the wrist. We report on a 52-year-old male who presented with methicillin-resistant Staphylococcus aureus (MRSA) arthritis of the wrist following a severe crush injury. We performed on this patient wrist arthrodesis using curettage of the lesion and vascularized fibular transfer with comparatively good results. Arthrodesis of the wrist using vascularized fibular transfer is useful for reconstructing bone destruction or defects accompanying intractable arthritis of the wrist such as caused by MRSA arthritis.
Assuntos
Artrite Infecciosa/cirurgia , Artrodese/métodos , Fíbula/transplante , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/cirurgia , Articulação do Punho/microbiologia , Artrite Infecciosa/tratamento farmacológico , Humanos , Masculino , Resistência a Meticilina , Pessoa de Meia-Idade , Radiografia , Articulação do Punho/diagnóstico por imagemRESUMO
Epidermolysis bullosa simplex associated with late onset of muscular dystrophy has been found to show defective expression of plectin, an intracytoplasmic protein in hemidesmosomes. In this report, we examined ability of cell-to-matrix attachment of cultured keratinocytes derived from a case with this disease by various cell biological methods, and compared it to that of normal keratinocytes. In cell adhesion assay, the patient keratinocytes showed more prominent short-time cell adhesion than normal keratinocytes. In contrast, the patient keratinocytes could be detached much easier than normal keratinocytes in cell detachment assay by treatment with dispase. In phagokinetic track assay, no apparent difference of cell migration was observed between the patient and normal keratinocytes. These results indicate that plectin-deficiency may up-regulate short-term cell contact and reduce stable cell-matrix adhesion at the epidermal basement membrane zone.
Assuntos
Epidermólise Bolhosa Simples/patologia , Proteínas de Filamentos Intermediários/deficiência , Queratinócitos/patologia , Adulto , Adesão Celular/genética , Células Cultivadas , Epidermólise Bolhosa Simples/genética , Epidermólise Bolhosa Simples/metabolismo , Humanos , Proteínas de Filamentos Intermediários/genética , Queratinócitos/fisiologia , Masculino , Mutação , PlectinaRESUMO
We reported a case of small cell carcinoma of the lung with metastasis to the iris during a stage of complete remission obtained with chemotherapy and radiation therapy. The patient was a 55-year-old man hospitalized for hoarseness and abnormal chest radiographs in August 1996. Small cell carcinoma of the lung had been diagnosed, and the stage was limited disease. Treatment consisted of 3 cycles of chemotherapy with cisplatin and etoposide, together with radiation therapy. The patient achieved complete remission and was discharged. In mid-December, he visited an eye clinic with the complaints of blurred vision and congestion in the right eye. Metastatic tumor of the iris was diagnosed. At that time, neither local recurrence of the lung cancer nor metastasis to other organs were observed. The patient was treated with cisplatin and etoposide again, resulting in a reduction of the iris tumor's size. After chemotherapy, the right eye was treated with electron irradiation, and the iris tumor and other clinical signs almost entirely disappeared. The patient retained normal vision during the clinical course.
Assuntos
Carcinoma de Células Pequenas/secundário , Neoplasias da Íris/secundário , Neoplasias Pulmonares/patologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma de Células Pequenas/terapia , Cisplatino/administração & dosagem , Etoposídeo/administração & dosagem , Humanos , Neoplasias da Íris/terapia , Neoplasias Pulmonares/terapia , Masculino , Pessoa de Meia-Idade , Radioterapia Adjuvante , Resultado do TratamentoRESUMO
A 46-year-old man presented with swallowing difficulty and dyspnea when in the supine position. Chest X-ray and computed tomographic (CT) films disclosed left pleural effusion and a tumor shadow extending invasively from superior to anterior mediastinum around the heart and large arteries. These observations called for a differential diagnosis from malignant lymphoma, invasive thymoma, and small cell carcinoma. Bronchofiberscopy and percutaneous tumor biopsy were performed, but the findings were inconclusive. Thoracoscopic biopsy yielded a diagnosis of sarcoidosis. No extrathoracic lesions were detected. Corticosteroid therapy (30 mg/day of prednisolone) was started. After 6 months of treatment (7.5 mg/day of prednisolone), the tumor shadow was reduced in size and the patient's swallowing difficulty and dyspnea subsided. This was a rare case of sarcoidosis extending invasively around the heart and large arteries, and that needed to be differentiated from mediastinal tumor. Thoracoscopic biopsy should be actively enlisted as a diagnostic procedure in difficult cases of this kind.
Assuntos
Doenças do Mediastino/diagnóstico , Neoplasias do Mediastino/diagnóstico , Sarcoidose/diagnóstico , Biópsia/métodos , Diagnóstico Diferencial , Humanos , Masculino , Pessoa de Meia-Idade , Cirurgia Torácica VídeoassistidaRESUMO
BACKGROUND: Epidermolysis bullosa simplex associated with muscular dystrophy (EBS-MD; OMIM# 226670) is an autosomal recessive disorder caused by genetic defects in the plectin gene. Because EBS-MD is relatively rare, and gene defects have been elucidated only in a limited number of patients, the precise phenotype-genotype correlations have not yet been fully elucidated. OBJECTIVE: The purpose of this study was to define clinical features of EBS-MD and to clarify its phenotype-genotype correlations. METHODS: Clinical, ultrastructural, immunohistochemical, and molecular features of 4 unrelated Japanese patients with EBS-MD were recorded. In addition, 6 cases with defined plectin gene mutations reported in the literature were reviewed. RESULTS: In skin of the EBS-MD patients, the blister formation always occurs just above the hemidesmosomes, and expression of plectin is absent or markedly reduced in all cases examined. All 10 patients, including 6 cases in the literature, showed generalized blistering at birth or soon thereafter, and experienced nail deformities. In addition, decayed teeth (5 cases), urethral strictures (3), mild palmoplantar hyperkeratosis (2), infantile respiratory complications (2), alopecia (1), and laryngeal webs (1) were present. All 8 patients who were older than 9 years demonstrated considerable muscle weakness, and the majority of them ended up being wheelchair bound. Among the 10 patients, 7 were products of consanguineous marriage, 9 have premature termination codon (PTC) mutations in both alleles of the plectin gene, and 7 cases were homozygous for the mutation. One patient who is homozygous for a 2719del9 in-frame deletion mutation that resulted in elimination of 3 amino acids, QEA, could still walk at the age of 46 and showed milder clinical severity. CONCLUSION: EBS-MD reveals clinical features not only characteristic of EBS and MD, but also other manifestations including urethral, dental, and respiratory complications. The majority of patients are products of consanguineous marriage and have homozygous plectin gene mutations. Whereas patients with PTC mutations in both alleles typically showed severe clinical features of EBS-MD and ended up being wheelchair bound, a homozygous patient for an in-frame deletion mutation showed positive, yet attenuated, plectin expression and milder clinical phenotype. Thus plectin immunofluorescence, combined with identification of the underlying plectin mutations, is of value in predicting the severity of the muscle involvement that occurs later in life of patients with EBS-MD.
Assuntos
Epidermólise Bolhosa Simples/complicações , Distrofias Musculares/complicações , Criança , Epidermólise Bolhosa Simples/genética , Feminino , Imunofluorescência , Genótipo , Humanos , Imuno-Histoquímica , Proteínas de Filamentos Intermediários/genética , Pessoa de Meia-Idade , Distrofias Musculares/genética , Mutação , Fenótipo , PlectinaRESUMO
A 21-year-old man was admitted to our hospital because chest X-ray films disclosed infiltrative shadows indicative of Mycoplasma pneumonia. He experienced fever and dry cough for 2 weeks. Chest X-ray findings showed ground-glass shadows in the lower fields of both lungs. The patient was not responsive to antibiotic therapy (PAPM/BP 1 g/day and CLDM 1,200 mg/day); dyspnea advanced rapidly and spikes of fever persisted. On hospital day 5, chest computed tomographic (CT) films disclosed interstitial shadows in all lower lung fields with dense infiltration. A transbronchial lung biopsy (TBLB) was performed on day 7, and TBLB specimens demonstrated infiltration of mononuclear cells in alveolar septa and organizing exudate in alveolar ducts with polypoid granulation tissue. Bronchoalveolar lavage fluid findings revealed an increase in the total cell count and the percentage of lymphocytes. The CD 4/CD 8 ratio was normal. The findings of other laboratory tests ruled out drug-induced lung disease, infectious disease, and collagen disease. Idiopathic bronchiolitis obliterans with organizing pneumonia (BOOP) was diagnosed. Corticosteroid therapy (methyl prednisolone: 500 mg/day) was started. After 2 weeks of treatment (prednisolone: 30 mg/day), the dyspnea and fever disappeared. Chest CT films showed that the interstitial shadows had largely resolved, but that a large cystic lesion had formed rapidly in the right lower lung field (right S 6). To the best of our knowledge, no cases of BOOP complicated by cystic lesions in the healing stage have been reported to date. We speculated that polypoid granulation in a bronchiole had given rise to a check-valve mechanism. After 2 months of treatment (prednisolone: 15 mg/day), the cystic lesion disappeared. We concluded that the bronchiolar lesion of polypoid granulation had resolved in response to therapy, thus facilitating air-way communication and the release of air from the cyst.
Assuntos
Pneumonia em Organização Criptogênica/complicações , Cistos/complicações , Pneumopatias/complicações , Adulto , Pneumonia em Organização Criptogênica/diagnóstico por imagem , Cistos/diagnóstico por imagem , Humanos , Pneumopatias/diagnóstico por imagem , Masculino , RadiografiaAssuntos
Dermatomicoses/diagnóstico , Dermatoses da Mão/diagnóstico , Pseudallescheria/isolamento & purificação , Idoso , Antifúngicos/uso terapêutico , Biópsia por Agulha , Dermatomicoses/tratamento farmacológico , Dermatomicoses/microbiologia , Dermatomicoses/patologia , Diagnóstico Diferencial , Fluconazol/uso terapêutico , Seguimentos , Dermatoses da Mão/tratamento farmacológico , Dermatoses da Mão/microbiologia , Dermatoses da Mão/patologia , Humanos , Masculino , Microscopia Eletrônica de Varredura , Pseudallescheria/ultraestruturaRESUMO
Home management for cancer-related pain in a terminal cancer patient is usually performed by oral or intrarectal administration with analgesics. If such treatment fails to reduce cancer-related pain, we have to treat them with epidural, intravenous or subcutaneous injection of these agents in some cases. We encountered a terminal lung cancer patient with metastatic bone cancer who was treated with continuous subcutaneous injection of morphine hydrochloride at home. As a result, the patient's quality of life has been remarkably improved. The management of cancer-related pain with analgesics and terminal care at home are discussed.
Assuntos
Neoplasias Ósseas/fisiopatologia , Neoplasias Ósseas/secundário , Serviços Hospitalares de Assistência Domiciliar , Neoplasias Pulmonares/patologia , Morfina/administração & dosagem , Dor Intratável/tratamento farmacológico , Assistência Terminal , Idoso , Humanos , Injeções Subcutâneas , MasculinoRESUMO
BACKGROUND: Bullous pemphigoid is an autoimmune bullous disease with circulating anti-basement membrane zone antibodies, and it commonly affects elderly individuals; however, the reasons for the late onset of the disease are unclear. DESIGN: The anti-basement membrane zone antibodies in serum samples from normal elderly subjects were compared with those in serum samples from normal young subjects. PARTICIPANTS: Serum samples from 32 elderly and 28 young normal individuals and 10 patients with bullous pemphigoid were used. INTERVENTIONS: Indirect immunofluorescence against guinea pig esophagus or human salt-split epidermis and immunoblotting against human and guinea pig epidermis were performed. RESULTS: Serum samples from young individuals were devoid of anti-basement membrane zone antibodies against guinea pig esophagus and human salt-split epidermis. Among 32 serum samples from elderly patients, 6 cases (19%) were positive for anti-basement membrane zone antibody for guinea pig esophagus, and in those the titers were 10 in 3 cases and 40, 80, and 320 in the others. One case was positive against human split epidermis at a titer of 10. An immunoblotting analysis showed that the antigenicity of the 230-kd and 180-kd bullous pemphigoid antigen from guinea pig epidermal extract was similar to that of human epidermal extract; however, the molecular weight was slightly different. The 4 cases of elderly serum that recognized guinea pig esophagus basement membrane zone showed positivity with the 230-kd peptide in the guinea pig epidermal extract; however, they were negative with the human epidermal extracts. Direct immunofluorescence observation of these cases showed that deposition of IgG or C3 was not present in cryostat sections from flexor arm surfaces. CONCLUSIONS: The serum samples from elderly subjects possessed a relatively high incidence of anti-basement membrane zone antibodies detectable with guinea pig esophagus as substrate. This observation of a specific immune defect in elderly individuals might explain why they are more susceptible to developing bullous pemphigoid.
Assuntos
Envelhecimento/imunologia , Autoanticorpos/análise , Membrana Basal/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Técnica Direta de Fluorescência para Anticorpo , Técnica Indireta de Fluorescência para Anticorpo , Cobaias , Humanos , Pessoa de Meia-Idade , Penfigoide Bolhoso/imunologia , Valores de ReferênciaRESUMO
In a distinct autosomal recessive variant of epidermolysis bullosa, EB-MD, life-long skin blistering is associated with late-onset muscular dystrophy of unknown etiology. Electron microscopy of these patients' skin suggests that tissue separation occurs intracellularly at the level of the hemidesmosomal inner plaque, which contains plectin, a high molecular weight cytoskeletal associated protein, also expressed in the sarcolemma of the muscle. In this study, we report two patients with EB-MD, each with a homozygous deletion mutation in the plectin gene, PLEC1. In the first case, the proband and her similarly affected sister had a homozygous 9 bp deletion mutation, designated as 2719de19, which resulted in elimination of three amino acids, QEA, in a sequence of 23 amino acids entirely conserved between the mouse and human sequences. The proband in the second family demonstrated a single nucleotide deletion at position 5866, designated as 5866delC, which resulted in frameshift and a premature termination codon for translation 16 bp downstream from the site of deletion. The absence of plectin in the hemidesmosomes, as reflected by negative immunofluorescence with an anti-plectin antibody (HD-1), associated with fragility of basal keratinocytes, implicates plectin as critical for binding of intermediate keratin filament network to hemidesmosomal complexes. The function of plectin as a putative attachment protein also in the muscle would explain the clinical phenotype consisting of cutaneous fragility and muscular dystrophy in EB-MD.
Assuntos
Epidermólise Bolhosa Simples/genética , Proteínas de Filamentos Intermediários/genética , Distrofias Musculares/genética , Animais , Epidermólise Bolhosa Simples/metabolismo , Epidermólise Bolhosa Simples/fisiopatologia , Feminino , Deleção de Genes , Homozigoto , Humanos , Masculino , Camundongos , Dados de Sequência Molecular , Linhagem , PlectinaRESUMO
FK506, a new immunosuppressive agent, is effective in treating patients with psoriasis. A major feature of psoriasis vulgaris is the hyperproliferation of keratinocytes together with inflammation. To determine the effect of FK506 or cyclosporin A (CsA) on the keratinocyte cell cycle, flow cytometry and the growth factor free normal human keratinocyte-arrested system were used to assess keratinocyte proliferation. FK506 and CsA inhibit keratinocyte proliferation induced by EGF, TGF-alpha or IL-6. The antiproliferative effects of FK506 and CsA directly correlated with blockade of the keratinocyte cell cycle at the G0/G1 phases. These findings might indicate that the effects of FK506 and CsA on proliferation of cultured normal human keratinocytes are probably related to direct effects on growth regulation of keratinocytes via EGF, TGF-alpha or IL-6 stimulation.
Assuntos
Ciclosporina/farmacologia , Imunossupressores/farmacologia , Queratinócitos/efeitos dos fármacos , Tacrolimo/farmacologia , Divisão Celular/efeitos dos fármacos , Linhagem Celular , DNA/biossíntese , Fator de Crescimento Epidérmico/farmacologia , Fase G1/efeitos dos fármacos , Humanos , Interleucina-6/farmacologia , Queratinócitos/citologia , Queratinócitos/metabolismo , Fase de Repouso do Ciclo Celular/efeitos dos fármacos , Fator de Crescimento Transformador beta/farmacologiaAssuntos
Apoptose , Carcinoma Basocelular/genética , Carcinoma Basocelular/patologia , DNA de Neoplasias/genética , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Animais , Técnicas Genéticas , Humanos , Masculino , Invasividade Neoplásica , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Orquiectomia , Próstata/citologia , Próstata/fisiologia , Ratos , Ratos WistarRESUMO
Hair follicle growth is thought to be regulated by a complex interplay of stimulatory and inhibitory signals. From among such signals, we examined the effects of transforming growth factor beta 1 (TGF beta 1) on the murine hair growth cycle. Quantitation of TGF beta 1 per wet tissue weight was performed at different stages of the hair cycle. TGF beta 1 was extracted with 10 mM HCl, and its level was measured with ELISA. The level of TGF beta 1 did not change markedly during the hair cycle. Immunohistochemical observations revealed that hair follicle cells and epidermal cells were negative for TGF beta 1 at all stages of the hair cycle. Mast cells in the dermis were positive throughout the cycle. To confirm whether the proliferation of hair follicles. Our observations suggest that TGF beta 1 is at least partly responsible for regulating hair follicles as a negative growth factor.
Assuntos
Folículo Piloso/efeitos dos fármacos , Mastócitos/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Ensaio de Imunoadsorção Enzimática , Folículo Piloso/crescimento & desenvolvimento , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos , Fator de Crescimento Transformador beta/farmacologiaRESUMO
In the previous study, we demonstrated apoptotic cells in basal cell carcinomas by means of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL). Positive staining cells were present at the peripheries and centers of the nests. The number of positive cells increased in cases showing deep infiltration. We also stained other epidermal neoplasms using this modified TUNEL method. In squamous cell carcinomas, many positive cells were observed in poorly differentiated types, but only a few cells were stained in well-differentiated types. TUNEL positive cells were rare in Bowen's disease. No positive cells were seen in seborrheic keratosis or verruca vulgaris. Our studies showed that apoptotic cells were more common in rapid growth neoplasms than in slow growth neoplasms.
Assuntos
Apoptose , Doença de Bowen/patologia , Carcinoma de Células Escamosas/patologia , Dano ao DNA , Neoplasias Cutâneas/patologia , DNA Nucleotidilexotransferase , DNA de Neoplasias/análise , Eletroforese , Humanos , Ceratose Seborreica/patologia , Dermatopatias/patologia , Coloração e Rotulagem/métodos , Verrugas/patologiaRESUMO
Epidermolysis bullosa is a group of inherited blistering diseases classified into three main sub-groups on the basis of the level of cleavage within the skin. In dominant dystrophic epidermolysis bullosa, characterized by cleavage below the basal lamina, two variants can be distinguished by the presence (Pasini form) or absence (Cockayne-Touraine form) of albo-papuloid lesions. The present study was undertaken to investigate the glycosaminoglycan chains of proteoglycans in the albo-papuloid lesions of a patient with the Pasini form, using histochemical and immunohistochemical methods. Histological examination revealed no dermo-epidermal separation. In the dermis, the papillary and subpapillary layers were slightly homogeneous, and exhibited a strong affinity towards alcian blue, which was abolished by treatment with chondroitinase ABC or in the presence of MgCl2 0.3M, but was resistant to digestion with streptomyces hyaluronidase. The papillary and subpapillary layers were intensely stained with a monoclonal antibody against small size proteoglycan with dermatan sulfate. These results suggest the presence of degraded dermatan sulfate proteoglycan in the papillary and subpapillary dermis of albo-papuloid lesions in the Pasini form of dystrophic epidermolysis bullosa.
Assuntos
Epidermólise Bolhosa Distrófica/metabolismo , Proteoglicanas/análise , Pele/química , Epidermólise Bolhosa Distrófica/patologia , Humanos , Imuno-HistoquímicaRESUMO
Recent advances in molecular biology have enabled the association of epidermolysis bullosa simplex (EBS) with point mutations of keratin 14 and/or keratin 5 genes to be established. We describe here the detection of point mutations in genomic DNA from formalin-fixed and paraffin-embedded sections from five cases of epidermolysis bullosa using the PCR amplification of specific alleles (PASA) method. In two of four cases of Köbner-type EBS a point mutation of helix 2b (384 Leu-Pro) was detected and in one case of Dowling-Meara-type EBS a mutation in helix 1a (125 Arg-Cys) was detected. The results of this study are consistent with previous reports and they demonstrate that the PASA method is a rapid and reproducible method for the detection of single-base changes and small deletions.
Assuntos
Alelos , Epidermólise Bolhosa Simples/classificação , Epidermólise Bolhosa Simples/genética , Queratinas/genética , Mutação Puntual , Reação em Cadeia da Polimerase/métodos , Sequência de Aminoácidos , Sequência de Bases , Primers do DNA , Humanos , Queratinas/química , Dados de Sequência Molecular , Estrutura Secundária de Proteína , Moldes GenéticosRESUMO
In the present study, we have examined whether or not the Paget cells in cases with dermal invasion or metastasis exhibit a different DNA distribution pattern to that seen in the epidermis in extramammary Paget's disease. The DNA contents of Paget cells from the epidermis and the tumor were determined by flow cytometry in two cases of extramammary Paget's disease with tumor formation, dermal invasion and metastases. The DNA histogram of Paget cells from the epidermis of two cases appeared to be close to the normal ploidy, whereas the histogram from the tumors was aneuploid in both cases. Our data suggest that the behavior of Paget cells in the epidermis is different from that in the tumor.
Assuntos
DNA/metabolismo , Neoplasias dos Genitais Masculinos/metabolismo , Doença de Paget Extramamária/metabolismo , Idoso , Idoso de 80 Anos ou mais , Núcleo Celular/metabolismo , DNA de Neoplasias/metabolismo , Epiderme/metabolismo , Citometria de Fluxo , Neoplasias dos Genitais Masculinos/patologia , Humanos , Imuno-Histoquímica , Masculino , Doença de Paget Extramamária/patologiaRESUMO
A 72-year-old woman had a 7-year history of a scaly red area on the right breast which had enlarged asymptomatically. A biopsy of the nipple was taken, and, following the finding of Paget's disease, a modified radical mastectomy was carried out. On histological examination of the entire breast specimen by serial sections, no evidence of an intraductal adenocarcinoma was found. Fractionation of Paget cells was performed from the epidermis. Stripped skin was treated with EDTA and trypsin, and epidermal cell suspensions were obtained. They were layered onto discontinuous Percoll gradients and centrifuged. Paget cells fell into three fractions with densities of 1.041, 1.058, and 1.078. Electronmicroscopically, the purity of fractionated cells obtained by this method ranged from 55 to 74% with viabilities of from 70 to 90%. Microscopic analysis of the DNA content in these cells was performed. The DNA histogram was close to the normal ploidy. This may explain why the mammary Paget's disease lesions in this case enlarged rather slowly.
Assuntos
Neoplasias da Mama/patologia , DNA de Neoplasias/análise , Doença de Paget Mamária/patologia , Idoso , Neoplasias da Mama/química , Neoplasias da Mama/ultraestrutura , Feminino , Histocitoquímica , Humanos , Microscopia Eletrônica , Doença de Paget Mamária/química , Doença de Paget Mamária/ultraestruturaRESUMO
We report a 19-year-old woman with typical clinical histological and immunofluorescence features of bullous pemphigoid. By immunoblotting, the serum was shown to detect antigens at 240 and 138 kDa. Elevated serum IgE levels were present, and there was a marked peripheral blood eosinophilia. The degree of eosinophilia correlated with small changes in the severity of the skin lesions, and the IgE level showed correlation with marked changes in severity. Another unusual feature was an exacerbation of her disease at the time of her first menses after the onset of the bullous pemphigoid.
Assuntos
Antígenos/análise , Penfigoide Bolhoso/imunologia , Adulto , Western Blotting , Eosinofilia/imunologia , Feminino , Humanos , Imunoglobulina E/análise , Peso Molecular , Penfigoide Bolhoso/patologiaRESUMO
In situ hybridization has been used for the detection of viral and bacterial mRNA and DNA which has infected tissue sections, but the sensitivity of this method is very limited. In this paper, a new, sensitive and simple non-isotopic in situ hybridization method in combination with a polymerase chain reaction (PCR) technique is described, in which the genomic or viral DNA can be amplified directly on formalin-fixed and paraffin-embedded tissue sections. As low as two copies of target DNA per cell can be detected by this in situ hybridization after in situ PCR. The essential modification concerns a two step cycling PCR procedure that was applied using single primer pairs. The first step, annealing, was achieved at a low temperature for a relatively long time. The second step was performed at a high temperature for a short period of time. This procedure greatly decreased the non-specific amplification of DNA and increased the sensitivity of detection.